The overall goal of this research is to examine the initial molecular interactions that are a critical first step in determining the ability of a bunyavirus to infect ist mosquito vector. Bunyaviruses are a significant cause of disease in humans and in domestic animals worldwide. Most bunyaviruses are transmitted to humans and animals by mosquitoes. They will use La Crosse encephalitis virus (LACV) and its natural mosquito vector Aedes triseriatus for these studies. They will examine the role viral glycoproteins play in the binding of virus to mosquito midgut, in the infection of Ae. triseriatus by LACV, and in the specificity of LACV for selected mosquito species. They will generate infectious LACV entirely from cloned cDNA using a RNA polymerase 1-based system that was recently used to reverse engineer influenza viruses. They will introduce specific mutatuions and chimeric segments and genes into LACV. Based on a significant body of work in their laboratory, they hypothesize that the G2 envelope protein of La Crosse virus is critical for virus binding to the cells of the mosquito midgut, and thus critical for mosquito infection. They will determine the protein sequences of G2 that are required for biding of La Crosse virus to the midgut cells of its natural mosquito vector, and to another permissive mosquito species. They will determine if vector specificity, in part, is mediated by pepetide sequences in the viral glycoproteins and if these sequences are the same that determine virus binding to midgut cells of competent vector mosquitoes. The development of efficient techniques to generate bunyaviruses containing specific mutations will permit in vivo investigations of individual gene functions in mosquito infections and in mammalian pathogenesis